Immunohistochemical localization of choline acetyltransferase during development and in Chats mutants of Drosophila melanogaster.

The distribution of choline acetyltransferase (CAT) in the nervous system of Drosophila melanogaster was determined by indirect immunohistochemical procedures using a monoclonal antibody specific to the enzyme. Immunoreactivity was first detected in the nervous system of 16 hr embryos, and increased considerably by the end of embryogenesis. Neuropil was preferentially stained, though cell bodies could also be observed. Staining was prominent in the CNS of all 3 larval instars but decreased substantially during the mid-pupal stage. Prior to eclosion, the level of immunoreactivity increased and the adult staining pattern became discernible. In the adult brain, staining was extensive, with numerous structures, such as the optic lobes and mushroom bodies, staining strongly. The adult thoracic ganglia were also moderately immunoreactive. These results imply a wide distribution of cholinergic neurons in the CNS of Drosophila. Immunoreactivity was also determined for 2 temperature-sensitive CAT mutants, Chats1 and Chats2. These files exhibit reduced CAT activity at permissive temperature, 18 degrees C, which eventually falls to undetectable levels after incubation at nonpermissive temperature, 30 degrees C. Chats2 mutants, after incubation at either 18 or 30 degrees C displayed virtually no staining. This result indicated that the immunoreactivity observed in wild-type flies was specifically associated with the enzyme encoded by the Cha gene. The intensity of staining in Chats1 mutants incubated at 18 degrees C appeared greater than in control flies, even though CAT enzyme activity in Chats1 is lower. This suggests that the enzyme molecule itself is structurally altered in Chats1 mutants. After incubation at 30 degrees C, staining in Chats1 flies decreased but did not disappear.